Apparent Digestibility of Protein and Amino Acids in Broiler Chickens Fed a Corn-Soybean Diet Supplemented with Microbial Phytase

Transcription

1 Apparent Digestibility of Protein and Amino Acids in Broiler Chickens Fed a Corn-Soybean Supplemented with Microbial S. SEBASTIAN, S. P. TOUCHBURN,1 E. R. CHAVEZ, and P. C. LAGUË Department of Animal Science, Macdonald Campus of McGill University, 21,111 Lakeshore Road, Ste. Anne de Belleuve, Quebéc, Canada, H9X 3V9 ABSTRACT The effect of microbial phytase supplementation on CP and amino acid (AA) digestibility was investigated in a 28-d trial using 360 sexed, day-old broiler chickens fed corn-soybean meal diets. The experimental design was a completely randomized one with a 3 2 factorial arrangement of treatments. The variables included P and Ca levels and phytase: P and Ca levels were: normal P-normal Ca [0.45% available P (P a ), 1.0% Ca], low P-normal Ca (0.35% P a, 1.0% Ca), and low P-low Ca (0.35 P a and 0.6% Ca); and phytase at 0 and 600 U/kg diet. supplementation increased body weight (P < 0.014) and feed (P < 0.004) at 19 d in male chickens; in females, phytase increased (P < 0.012) only body weight at 19 d. The low P- normal Ca diet reduced (P < 0.05) feed and body weight in both sexes at 7, 14, and 19 d, compared to the normal P-normal Ca diet; the reduction of Ca in the low P diet prevented the above depression, resulting in body weight and feed to a level comparable to that of the normal P-normal Ca diet. Microbial phytase supplementation had no effect (P < 0.065) on the apparent ileal digestibility (AID) of CP or any AA except Met and Phe in male broiler chickens. In females, adding phytase increased the AID of all AA except Lys, Met, Phe, and Pro. The low P-normal Ca diet reduced (P < 0.05) the AID of Phe, Asp, and Ser in male chickens and reduced the AID of all the AA except Met and Pro in females compared to the normal P-normal Ca diet. The reduction of Ca in the low P diet prevented the depression of the AID of the AA caused by the low P- normal Ca diet, resulting in AID of AA having a level comparable to that of the normal P-normal Ca diet in both sexes. supplementation did not have any effect (P > 0.05) on apparent fecal digestibility (AFD) of CP or any of the AA in male chickens; however, in female chickens it increased the AFD of Thr, Asp, Glu, and Ser. In summary, phytase supplementation increased growth performance in both sexes; increased AID and AFD of most of the AA, particularly in female chickens. The optimum growth performance and AA digestibilities were obtained with the lowest input of resources, in the low P-low Ca diet supplemented with microbial phytase. (Key words: broiler, phytase, calcium levels, crude protein, amino acids) 1997 Poultry Science 76: INTRODUCTION Phytic acid, myo-inositol phosphorylated on all of its six hydroxyl groups, can ionically bind minerals and proteins in aqueous medium (de Rham and Jost, 1979). The interactions among phytic acid, minerals, and protein appear to be primarily responsible for the adverse nutritional effects of a high-phytate diet (Cheryan, 1980). Phytic acid has the ability to bind protein at acidic, alkaline, and neutral ph (Anderson, 1985). The interaction between phytic acid and protein leads to decreased solubility of protein and eventually reduces its utilization (Cheryan, 1980). It is well documented that microbial phytase supplementation enhances the phytic acid hydrolysis and Received for publication April 7, Accepted for publication August 14, To whom correspondence should be addressed. increases the availability of minerals bound to the phytic acid. Theoretically, phytase supplementation must also be able to release the phytate-bound protein for utilization. It has been reported that phytase supplementation improved N digestibility in pigs (Yi et al., 1994), and N retention in broiler chickens (Farrell et al., 1993) and laying hens (Van der Klis and Versteegh, 1991). In contrast, Newkirk and Classen (1995) did not find any significant effect on CP digestibility by either semipurified phytase or crude phytase supplementation to broiler diets. In a recent study with female turkeys, Yi et al. (1996) showed that phytase supplementation enhanced the ileal digestibility of N and AA. Mroz et al. (1994) reported that phytase supplementation enhanced the apparent digestibility of CP and AA except for cystine and Pro in pigs. There is no published data on the apparent ileal digestibility of CP and AA as influenced by supplementary microbial phytase in diets for broiler chickens. It has been shown that dietary Ca 1760

2 EFFECTS OF PHYTASE ON AMINO ACID DIGESTIBILITY 1761 and P levels affect the efficacy of microbial phytase on mineral utilization in broiler chickens (Sebastian et al., 1996a). The objective of this experiment was to investigate the efficacy of microbial phytase supplementation on apparent digestibility of protein and AA in broiler chickens fed either normal P-normal Ca or low P-normal Ca or low P-low Ca corn-soybean meal diets. MATERIALS AND METHODS Experimental Design A total of 360 sexed, day-old broiler chicks was purchased from a commercial hatchery,2 wing-banded, and weighed individually. Ten birds were assigned to each of 36 pens (18 pens of each sex) and housed in thermostatically controlled Petersime3 battery brooders with raised wire floors. All birds were exposed to continuous fluorescent light. and water were provided for ad libitum consumption throughout the 28-d trial. Each of six treatments was replicated three times per sex (n = 3). The experimental design was completely randomized with a 3 2 factorial arrangement of treatments. The variables included P and Ca levels and phytase.4 Phosphorus and Ca levels were: normal P- normal Ca [0.45% available P (P a ), 1.0% Ca], low P-normal Ca (0.35% P a, 1.0% Ca), and low P-low Ca (0.35% P a, 0.6% Ca); and phytase at 0 and 600 U/kg diet. The percentage composition of the basal diets is shown in Table 1 and the AA analysis of the diets is shown in Table 2. Individual body weights of chickens and group feed consumption data were recorded on Days 7, 14, and 19. All birds were cared for according to the Canadian Council on Animal Care guidelines (CCAC, 1993). TABLE 1. Composition of experimental diets 1 Normal P- Low P- Low P- Ingredients and analyses normal Ca normal Ca low Ca Corn, ground Soybean meal (48% CP) Fish meal (65% CP) AV-fat Calcium carbonate (32% Ca) Dicalcium phosphate (25% Ca; 19% P) Salt Vitamin-mineral mixture DL-Methionine L-Lysine HCl Analyzed composition Ca Total P CP Calculated composition ME, kcal/kg 3,150 3,174 3,197 Available P Lysine Met + Cys Calculated to meet or exceed the requirements of broiler starter diet (NRC, 1994). 2Animal-vegetable fat blend. 3Vitamin-mineral premix supplied the following per kilogram of diet: Ca, 1,000 mg; P, 450 mg; magnesium, 25 mg; sodium chloride, 550 mg; selenium, 0.04 mg; iron, 25 mg; manganese, 22 mg; copper, 1.6 mg; zinc, 16 mg; iodine, 0.14 mg; retinyl acetate, 925 IU; cholecalciferol, 275 IU; dla-tocopherol acetate, 5 IU, choline, 40.8 mg; menadione sodium bisulfite, 0.19 mg; riboflavin, 1.9 mg; vitamin B 12, 2.2 mg; niacin, 8.1 mg; pantothenic acid, 2.2 mg; biotin, mg; folic acid, 0.16 mg; thiamin, 0.5 mg; pyridoxine, 0.9 mg; salinomycin sodium, 7.5 mg. (%) Sample Collection and Chemical Analysis The digestibilities of protein and AA were measured at 28 d. During this period, to attain uniformity in feed and digesta content, the feed troughs were removed for 30 min, then replaced for 2 h before the birds were taken out for the measurements. Four birds were randomly selected from each replicate at 28 d for the measurement of ileal apparent digestibility and apparent total digestive tract digestibility of protein and AA. After euthanasia with an intravenous injection of pentobarbitone, each bird was immediately dissected and the ileum located, defined as extending from Meckel s diverticulum to the ileo-cecal junction; the distal 40 mm of the ileum were tied off and excised. This segment was bisected transversely and its contents were gently squeezed out into a plastic cup. A 2René Poirier Ltée., St. Félix-de-Valois, QC, Canada, J0K 2M0. 3Gettysburg, OH Natuphos 1000, BASF Canada, Inc., Ville St. Laurent, QC, Canada H4T 1Y4. TABLE 2. Amino acid analysis of experimental diets 1 Normal P- Low P- Low P- Amino acids normal Ca normal Ca low Ca Essential Arg His Ile Leu Lys Met Phe Thr Val Nonessential Ala Asp Glu Gly Pro Ser Tyr Total amino acid The feed samples, in duplicate, were hydrolyzed with 6N HCl at 110 C and analyzed for amino acid content by HPLC using the Pico-Tag method from Waters chromatography systems. (%)

3 1762 SEBASTIAN ET AL. TABLE 3. The effect of phytase supplementation on feed, body weight, and feed to ratio of male broiler chickens fed corn-soybean diets for 19 d 0 to 7 d 0 to 14 d 0 to 19 d : (U/kg) (g) (g:g) (g) (g:g) (g) (g:g) Normal P-normal Ca Normal P-normal Ca Low P-normal Ca Low P-normal Ca Low P-low Ca Low P-low Ca SEM Normal P-normal Ca 130 ab 103 a a 335 a 1.34 b 722 a 557 a 1.30 Low P-normal Ca 124 b 95 b b 281 b 1.47 a 624 b 494 b 1.27 Low P-low Ca 132 a 101 ab ab 297 b 1.47 a 684 a 542 a b 513 b a 549 a 1.29 a,bmeans within columns with no common superscript differ significantly (P < 0.05). : : digesta sample from the rectum was also obtained to determine the apparent fecal digestibility. The digesta of two birds out of four were pooled separately in order to have more replicates for analysis. The pooled samples were freeze-dried, ground through 1-mm mesh screen, and immediately prepared for the analysis of CP and AA. Digesta and feed were hydrolyzed for 24 h with 6 N HCl at 110 C for the determination of AA by HPLC5 using the Pico-Tag system from Waters chromatography systems.6 Cystine and Trp, being partly destroyed under acid hydrolysis, were not determined. Nitrogen analysis was carried out with a Leco N analyzer.7 Chromic oxide was analyzed using a flame atomic absorption spectrophotometer8 after wet digestion with concentrated nitric acid and perchloric acid (Emerson, 1975). All samples were assayed in duplicate. Calculations and Statistical Analysis Apparent ileal and fecal digestibility coefficients of CP and AA were estimated by using 0.2% chromic oxide (Cr 2 O 3 ), at the expense of corn, as an indigestible marker. The following formula was used to calculate the digestibility coefficients: 5Waters Model 700 WISP Satellite; model 510 HPLC pump, Waters, Houston, TX Waters Chromatography Division, Millipore Corp., Milford, MA Model FP-428, Leco Corp., St. Joseph, MI Model 2380, Perkin Elmer, Norwalk, CT AD AA = [AA] digesta [AA] diet [Cr] diet [Cr] digesta where AD AA = apparent digestibilities of individual AA (percentage); [AA] digesta = AA concentration in the digesta; [AA] diet = AA concentration in the diet; [Cr] digesta = chromic oxide concentration in the digesta; [Cr] diet = chromic oxide concentration in the diet. The data on performance and apparent ileal and fecal digestibilities were analyzed using the General Linear Models procedure for analysis of variance (SAS Institute, 1990). Significant differences among treatment means were separated by Duncan s new multiple range test (Duncan, 1955) with a 5% level of probability. RESULTS Intake, Body Gain, and Efficiency The growth performance of male and female broiler chickens fed the various dietary treatments are shown in Tables 3 and 4, respectively. The main effect of microbial phytase (600 U/kg diet) indicates increased body weight (P < 0.014) and feed (P < 0.004) at 19 d in male chickens; however phytase supplementation did not have any significant (P > 0.05) effect on body weight or feed at either 7 or 14 d in female chickens, although phytase supplementation increased (P < 0.012) body

4 EFFECTS OF PHYTASE ON AMINO ACID DIGESTIBILITY 1763 TABLE 4. The effect of phytase supplementation on feed, body weight, and feed to ratio of female broiler chickens fed corn-soybean diets for 19 d 0 to 7 d 0 to 14 d 0 to 19 d : : : (U/kg) (g) (g:g) (g) (g:g) (g) (g:g) Normal P-normal Ca Normal P-normal Ca Low P-normal Ca Low P-normal Ca Low P-low Ca Low P-low Ca SEM Normal P-normal Ca 126 a 96 a a 304 a 1.42 b 673 a 509 a 1.33 Low P-normal Ca 120 b 89 b b 259 b 1.52 a 589 b 449 b 1.31 Low P-low Ca 129 a 98 a a 294 a 1.44 b 659 a 524 a b a 1.27 a,bmeans within columns with no common superscript differ significantly (P < 0.05). weight at 19 d in females (Table 4). had no influence (P > 0.05) on feed to ratio in either male or female chickens at 7, 14, or 19 d. The s in the diet caused significant effects on feed (P < 0.018) and body weight (P < 0.004) at 14 and 19 d in male chickens (Table 3) and on feed and body weight at 7, 14, and 19 d in female chickens (Table 4). The low P-normal Ca diet reduced (P < 0.05) feed, and body weight in both male and female chickens at 7, 14, and 19 d compared to the normal P-normal Ca and low P-low Ca diets with the exception of feed at 7 d in males. and body weight in both male and female chickens fed a low P-low Ca diet at 7, 14, and 19 d were comparable to that of the normal P-normal Ca diet except weight at 14 d in males. The interactions of by phytase were nonsignificant (P > 0.05) for any growth performance variables. The best performance overall after 19 d was obtained with the diet low P-low Ca supplemented with phytase. AID of CP and Essential Amino Acids The AID coefficients of CP and EAA of male and female chickens fed the various dietary treatments for 28 d are shown in Tables 5 and 6, respectively. ary treatments affected (P < to 0.042) the AID of CP and all of the EAA except His, Leu, Ile, Lys, and Val in male chickens (Table 5) and of all of the EAA except Met in female chickens (Table 6). The main effect of microbial phytase indicated that, in male chickens, phytase had no influence on the AID of CP (P < 0.069) and any AA except Met and Phe, which were reduced (P < 0.05) by phytase (Table 5). However, in female chickens, phytase supplementation significantly increased (P < 0.035) the AID of all of the EAA except Lys, Met, and Phe (Table 6). The had a significant effect on all of the EAA except Met in female chickens (Table 6), but had no significant effect on AID of any of the EAA except Phe (P < 0.007) in male chickens. The low P-normal Ca reduced (P < 0.05) the AID of all of the EAA except Met in female chickens and of Phe in male chickens compared to the normal P-normal Ca control diet. The low P-low Ca diet promoted the AID of EAA to a level comparable to that of the normal P-normal Ca diet in both sexes except for Phe and Thr in female chickens. The interaction of by phytase was significant for the AID of CP, His, Lys, Met, and Thr in male chickens; however, in female chickens, such interactions were nonsignificant (P > 0.05) for the AID of CP and all of the EAA. The AID of Nonessential Amino Acids The AID coefficients of NEAA of male and female chickens fed the various dietary treatments are shown in Tables 7 and 8, respectively. ary treatments affected (P < to 0.036) the AID of Gly, Pro, and Ser in male chickens (Table 7) and of all of the NEAA except Pro in female broiler chickens (Table 8). supplementation had no influence on AID of any of the NEAA in male chickens (Table 7); however, phytase supplementation increased (P < to 0.029) all of the NEAA except Pro in female chickens (Table 8). The had a significant

5 1764 SEBASTIAN ET AL. TABLE 5. Effect of phytase on apparent ileal digestibility of protein and essential amino acids in 28-d-old male chickens CP Arg His Ile Leu Lys Met Phe Thr Val Normal P-normal Ca ab ab ab 89.7 bc b 82.7 Normal P-normal Ca a a ab 91.0 abc a 85.8 Normal P-normal Ca a ab a 93.6 a b 83.3 Normal P-normal Ca b b b 87.8 c b 79.2 Low P-low Ca a ab a 91.5 ab ab 85.6 Low P-low Ca ab b ab 89.5 bc b 81.6 SEM Normal P-normal Ca a Low P-normal Ca b Low P-low Ca ab a 86.2 a b 83.6 b a cmeans within column with no common superscript differ significantly (P < 0.05). effect on Asp and Ser (P < 0.026) in male chickens and on all of the NEAA except Pro (P < to 0.045) in female chickens. The low P-normal Ca diet reduced (P < 0.05) the AID of Asp and Ser in male chickens and of all the NEAA except Pro in female chickens; however, the reduction of Ca in the low P diet increased the AID of NEAA, except Asp in both sexes, to a level comparable to that of the normal P-normal Ca diet in both sexes. The interaction of by phytase was significant (P < to 0.028) only for Ala, Gly, and Pro in male chickens; however, such interaction was nonsignificant (P > 0.05) for all of the NEAA in female chickens. supplementation to the low P-normal Ca diet reduced (P < 0.05) the AID of Gly and Pro in male chickens. TABLE 6. Effect of phytase on apparent ileal digestibility of protein and essential amino acids in 28-d-old female chickens CP Arg His Ile Leu Lys Met Phe Thr Val Normal P-normal Ca Normal P-normal Ca Normal P-normal Ca Normal P-normal Ca Low P-low Ca Low P-low Ca SEM Normal P-normal Ca a 87.1 a 82.4 a 89.9 a 90.1 a a 79.3 a 86.2 a Low P-normal Ca b 83.8 b 78.1 b 87.6 b 87.6 b b 74.4 b 82.6 b Low P-low Ca a 85.1 ab 81.3 a 89.3 ab 89.6 a b 78.5 b 85.1 ab b 84.2 b 79.0 b 88.0 b b 83.4 b a 86.5 a 82.1 a 89.9 a a 85.9 a a,bmeans within column with no common superscript differ significantly (P < 0.05).

6 EFFECTS OF PHYTASE ON AMINO ACID DIGESTIBILITY 1765 TABLE 7. Effect of phytase on apparent ileal digestibilities of nonessential amino acids in 28-d-old male chickens Ala Asp Glu Gly Pro Ser Tyr Normal P-normal Ca ab abc 88.7 bc Normal P-normal Ca a a 91.5 ab Low P-normal Ca a ab 93.9 a Low P-normal Ca b c 87.0 c Low P-low Ca a ab 92.4 ab Low P-low Ca ab bc 91.3 ab SEM Normal P-normal Ca a a 86.6 Low P-normal Ca b b 83.5 Low P-low Ca b ab a cmeans within column with no common superscript differ significantly (P < 0.05). The AFD of CP and EAA The apparent fecal digestibility coefficients of CP and EAA of female chickens fed the various dietary treatments are shown in Table 9. ary treatments had a significant effect (P < to 0.002) on the AFD of all of the EAA except Met in male chickens (data not presented); but in female chickens, dietary treatment was significant (P < to 0.019) only for Arg, His, Phe, and Thr (Table 9). The main effect of phytase indicates that phytase supplementation did not have any significant effect (P > 0.05) on AFD of any of the EAA in either sex except Thr in TABLE 8. Effect of phytase on apparent ileal digestibilities of nonessential amino acids in 28-d-old female chickens Ala Asp Glu Gly Pro Ser Tyr Normal P-normal Ca Normal P-normal Ca Low P-normal Ca Low P-normal Ca Low P-low Ca Low P-low Ca SEM Normal P-normal Ca 88.8 a 90.9 a 92.6 a 83.9 a a 88.2 a Low P-normal Ca 85.4 b 86.3 b 89.8 b 78.8 b b 84.9 b Low P-low Ca 87.8 a 86.0 b 91.4 a 81.9 a a 86.6 ab b 85.9 b 90.2 b 80.3 b b 85.2 b a 89.6 a 92.3 a 83.1 a a 88.0 a a,bmeans within column with no common superscript differ significantly (P < 0.05).

7 1766 SEBASTIAN ET AL. TABLE 9. Effect of phytase on apparent fecal digestibility of protein and essential amino acids in 28-d-old female chickens CP Arg His Ile Leu Lys Met Phe Thr Val Normal P-normal Ca Normal P-normal Ca Normal P-normal Ca Normal P-normal Ca Low P-low Ca Low P-low Ca SEM Normal P-normal Ca a 86.9 a a 80.1 a 86.4 a Low P-normal Ca b 80.8 b b 71.7 b 81.6 b Low P-low Ca a 84.4 a b 77.8 a 83.9 ab b a 84.8 a,bmeans within column with no common superscript differ significantly (P < 0.05). female chickens. The significantly affected (P < to 0.04) the AFD of all of the EAA in male chickens (data not presented) and of Arg, His, Phe, Thr, and Val in female chickens (Table 9). The low P-normal Ca diet reduced (P < to 0.03) the AFD of all of the EAA in male chickens (data not presented); in female chickens it reduced the AFD for Arg, His, Phe, Thr, and Val. The low P-low Ca diet resulted in (P < 0.05) AFD values of all the EAA comparable to that of the normal P-normal Ca diet in both sexes except for Phe in females. The interaction of P- Ca level by phytase was nonsignificant (P > 0.05) for the AFD of CP and for all of the EAA in both sexes. The AFD of NEAA The AFD coefficients for the NEAA of female chickens fed the various dietary treatments is shown in Table 10. ary treatments had a significant (P < to 0.05) effect on AFD of all of the NEAA in both sexes except Ala and Pro in female chickens. supplementation showed no significant (P > 0.05) effect on AFD of any of the NEAA in male chickens (data not presented); however in female chickens it produced a significant effect (P < 0.01 to 0.049) on AFD of Asp, Glu, and Ser (Table 10). The P-Ca levels also had significant influence (P < to 0.015) on AFD of all of the NEAA in both male and female chickens, except Pro in female chickens. The low P-normal Ca diet reduced (P < 0.05) AFD of all NEAA in both male and female chickens except Pro in female chickens. Lowering the Ca level in the low P-normal Ca diet increased (P < 0.05) AFD of all the NEAA, except Asp for females, affected by a low P-normal Ca diet to the levels similar to the normal P-normal Ca controls. The interaction of P-Ca level by phytase was nonsignificant (P > 0.05) for the AFD of any of the NEAA in both sexes. DISCUSSION The study presented herein shows the effects of microbial phytase supplementation at different P-Ca levels on AID and AFD of CP, EAA, and NEAA in both male and female chickens fed a corn-soybean meal diet for 28 d. In agreement with previous studies (Denbow et al, 1995; Kornegay et al., 1996; Sebastian et al., 1996b), phytase supplementation increased body weight and feed in both sexes at 19 d. However, phytase supplementation did not affect the feed to ratio because of the simultaneous increase in body weight and feed, an observation that agrees with those of Simons et al. (1990), Perney et al. (1993), Denbow et al. (1995), and Sebastian et al. (1996b). The absence of a significant interaction of by phytase in this study indicates that the influence of phytase on growth performance is independent of s in the diet. The observations of reduced feed and weight in chickens fed a low P-normal Ca diet may be due to: 1) inadequate supply of P in the diet to meet the P requirement, 2) reduction of appetite as a result of the low level of P, or 3) reduction in the solubility of mineral complexes as a consequence of increased ileal ph by the relatively high dietary Ca level (Shafey et al, 1991). These observations clearly indicate the importance of maintaining the ideal Ca:P ratio in broiler diets in order to obtained optimum growth performance. The low P-low Ca diet prevented the depression of feed and body weight observed in the low P-normal Ca diet;

8 EFFECTS OF PHYTASE ON AMINO ACID DIGESTIBILITY 1767 TABLE 10. Effect of phytase on apparent fecal digestibility of nonessential amino acids in 28-d-old female chickens Ala Asp Glu Gly Pro Ser Tyr Normal P-normal Ca Normal P-normal Ca Low P-normal Ca Low P-normal Ca Low P-low Ca Low P-low Ca SEM Normal P-normal Ca 88.5 a 90.6 a 92.0 a 82.1 a a 88.5 a Low P-normal Ca 83.3 b 84.3 b 88.6 b 74.3 b b 84.0 b Low P-low Ca 86.1 ab 84.0 b 90.4 ab 79.8 a a 85.8 ab b 89.4 b b a 91.3 a a 87.1 a,bmeans within column with no common superscript differ significantly (P < 0.05). in fact, the low P-low Ca diet increased the body weight and feed to a level comparable to that of the normal P-normal Ca diet. This study clearly revealed that phytase supplementation caused the optimum response (maximum output for the minimum input of resources) for growth performance when added to a low P-low Ca diet. To the authors knowledge, there are no published data on the AID coefficients of CP or AA as influenced by supplementary microbial phytase in diets of broiler chickens. The results of this experiment indicate that the addition of 600 U of phytase/kg diet increases AID of EAA as well as NEAA in female chickens; however, the magnitude of response to phytase in male chickens was very low. We don t have an obvious explanation of why phytase supplementation has more prominent effects on AA digestibilities in female than in male chickens. We are not aware of any published data indicating the effect of phytase supplementation on AA digestibilities in either male or female chickens that can be compared with our findings. Further investigations are needed to have a better understanding of this major discrepancy between female and male response. It is interesting to note that AID values of CP and all AA were higher in female than in male chickens. The observation, though inconsistent, of improved availability of AA by microbial phytase supports the notion that phytic acid binds the protein and AA and makes them unavailable (Cosgrove, 1980). Phytic acid does form complexes with proteins, the nature of which is governed by ph. Phytic acid is strongly negatively charged in the low ph range, whereas proteins are strongly positively charged; as a result, a phytate-protein complex can be formed (Saio et al., 1967). Although proteins and phytic acid have a net negative charge above ph 5, multivalent cations such as Ca, Mg, or Zn are thought to mediate the formation of phytate-protein complexes (Okubo et al., 1976). The formation of a phytate-protein complex can adversely affect certain functional properties, solubility, and digestibility of proteins (Reddy et al, 1982). Saio et al. (1967) showed that excess Ca ions interact with protein and phytate to further decrease the solubility of proteins. It has been demonstrated that phytate-protein complexes are less subject to proteolytic digestion than the same protein alone (Barre et al., 1956). When microbial phytase cleaves the ester bond to release P from phytate, it also frees proteins bound to phytate and increases the availability of protein and AA for absorption. The increase in the ileal digestibility of AA may also indicate that phytate-protein complexes were cleaved by microbial phytase to some extent. Phytate is also known to inhibit a number of digestive enzymes such as pepsin (Camus and LaPorte, 1976), and trypsin (Singh and Krikorian, 1982; Caldwell, 1992). These negative influences may also be partly responsible for the effects of phytate on protein utilization. Another possible reason for the improvement in AA digestibility is that phytase supplementation, by reducing phytic acid level in the diet, reduces the inhibitory effect of phytic acid on pepsin and trypsin, thereby increasing the digestibility of AA. In a recent study with female turkeys, Yi et al. (1996) showed that microbial phytase supplementation to a diet containing 0.45% nonphytate P improved ileal and true digestibilities of N and most of the EAA and NEAA at both 22.5 and 28.0% CP levels; at 0.60%

9 1768 nonphytate P level, phytase supplementation increased the ileal and true digestibilities of N and all of the AA at 22.5% CP level but not at the level of 28.0% CP. In contrast, a study with pigs showed that microbial phytase supplementation increased the AID of only Met and Arg, but did not influence the AID of other AA (Mroz et al., 1994). In the present study, the improvement in AID of AA by phytase supplementation was in a range of 1.8 (Arg) to 4.3 (Thr) percentage units, which is higher than that observed in a study dealing with female turkeys (Yi et al., 1996). We have no explanation for the decreased and increased digestibilities in the low P-normal Ca and the low P-low Ca diets, respectively. It may be possible that the former increases the ileal ph due to the presence of the relatively high Ca level compared to P in the diet (Shafey et al., 1991); at the consequently high ph, the excess Ca in the low P- normal Ca diet plays an important role in the formation of a phytate-ca-protein complex which is extremely insoluble (Saio et al., 1967) and thus reduces the availability of protein (and AA). The low P-low Ca diet prevents the situations of increased ileal ph and excess Ca, thereby reducing the formation of the insoluble complex. The range (76.8 to 91.8% and 76.2 to 90.6%) of average AID and AFD values of all AA reported in this study, are within the range reported previously in chickens (Payne et al., 1968; Green et al., 1987) and pigs (Mroz et al., 1994); however, the values reported by Yi et al., (1996) for AID in the turkey were consistently higher for all AA. The highest AID and AFD values were observed for Met and Pro among the EAA and NEAA, respectively, which agree with the results of Yi et al. (1996), who obtained the maximum AID value for Met in turkeys. The efficacy of phytase on the digestibility of AA in female chickens was further confirmed by the observation of increased AFD of Asp, Glu, Gly, Thr, and Ser in female but not in male chickens, emphasizing the need of further research to better understand these sex differences. It is not clear why phytase failed to show prominent effects on the AFD of the other AA in female chickens. It may be possible that the contribution of microbial protein and bacterial synthesis of some AA in the hind gut and ceca of chickens (Parsons et al., 1981) overshadows the effect of phytase. In contrast, Mroz et al. (1994) reported that microbial phytase supplementation in pigs enhanced the AFD of all AA except for cystine and Pro. As expected, AID of CP and all the AA was higher than AFD of the same in both male and female chickens, further indicating the potential contribution of microbes in the hind gut. These observations suggest that recovery of AA from the ileum represents a sensitive index of digestibility coefficient that is not confounded by cecal and intestinal bacteria. Contradictory to our observation, the results of Mroz et al (1994) in pigs show that AID values of all AA are lower than the AFD values. As observed in the case of AID, the AFD values of CP and all AA were numerically higher in female than male chickens. SEBASTIAN ET AL. In conclusion, the data obtained from this research show that dietary microbial phytase supplementation (600 U/kg diet) improved growth performance and AID of all AA except Lys, Met, Phe and Pro in female chickens. However, phytase had no influence on the AID of any of the AA except Met and Phe in male chickens; thus, the response to phytase supplementation may be sex dependent. supplementation increased the AFD of Asp, Glu, Thr, and Ser in female but not in male chickens. The growth performance and digestibility of AA observed in the low P-low Ca diet were comparable to those observed in the normal P- normal Ca diet at 28 d. In this study, phytase supplementation in the low P-low Ca diet showed optimum response for growth performance in both sexes, and for AID and AFD of AA, particularly in female chickens. ACKNOWLEDGMENTS The authors wish to thank BASF Canada, Inc., Ville St. Laurent, QC, for financial support of this study and for the phytase enzyme provided for this experiment and Zully Valencia, Luc Normand, Antoine Farhat, Sophie Lavallée, and Josée Chicoine for their assistance in sample collection and Denise Gaulin for assistance in lab analyses. REFERENCES Anderson, P. 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